Expression systems are utilized for producing useful gene products by genetic engineering depending on the purposes. In the expression systems, hosts for which techniques for their cultivation have been established such as microbial cells (Escherichia coli, Bacillus subtilis, yeast, etc.), animal cells, insect cells and plant cells, and promoters suitable for the hosts are used. Among these, an expression system in which Escherichia coli as a host and the lac promoter or a derivative thereof are used is one of the most commonly used systems because of the operational convenience.
However, the expression system in which the lac promoter or a derivative thereof is used has a drawback in that it is industrially disadvantageous because it requires induction of gene expression for expression of a gene product. For example, induction of gene expression from the lac promoter, the tac promoter or the like requires use of an expensive reagent, isopropyl-β-D-thiogalactopyranoside (IPTG). Therefore, such a system has a drawback in that it is disadvantageous to performance on an industrial scale.
An expression system in which a promoter derived from xylose operon and a bacterium of the genus Bacillus as a host are utilized is also used. However, the system is disadvantageous to performance on an industrial scale because it requires addition of xylose for the expression induction.
Expression vectors that utilize thermoinduction of the phage λ promoter are generally used.
However, overexpression of recombinant gene products by thermoinduction may be disadvantageous in the following points:
(a) difficulty in rapidly achieving shifting-up of a temperature;
(b) increased possibility of forming insoluble inclusion bodies due to a higher cultivation temperature; and
(c) induction of several proteases in Escherichia coli upon heat shock.
Bacillus subtilis is known to produce and secrete a number of catabolic enzymes such as amylases and proteases as a result of a stationary phase-specific response. If one could express a gene only during stationary phase after full growth of a host utilizing the stationary phase-specific expression mechanism, the burden on the host might be decreased, and an exogenous gene might be efficiently expressed. However, no gene expression technique in which such a mechanism is utilized has been established.
Thus, a technique that enables efficient expression without artificially inducing gene expression has been desired.